Glucose is the primary carbon and energy source for nearly all cells, so it is critical that they sense glucose and respond to it appropriately. The importance of this is especially apparent in mammals, which exhibit the severe pleiotropic phenotype called diabetes if they have even minor defects in glucose sensing. Glucose sensing and signaling is of special significance to yeast, because it largely determines the novel fermentative lifestyle of the organism. Our long-term goals are to understand how yeast cells sense glucose and generate intracellular signals, and how those signals are transduced to target proteins that affect cellular function. A major effect glucose has on yeast cells is alteration of gene expression. Glucose represses expression of genes not important to glucose-grown cells, and activates expression of genes in demand in cells growing on glucose. Two glucose signal transduction pathways responsible for these effects have recently come into focus. The glucose repression pathway includes a protein kinase (Snf1-Snf4) and a protein phosphatase (Reg1- Glc7) responsible for glucose activation of the Mig1 repressor. The glucose-induction pathway includes two glucose sensors in the membrane (Snf3 & Rgt2) that bind glucose and generate an intracellular signal that affects the ability of a ubiquitin ligase protein complex (SCFGrr1) to inhibit function of the Rgt1 repressor. To understand the mechanism of glucose induction (Aim 1), we need to learn a) how Rgt1 function is regulated by glucose, b) if and how the SCFGrr1 protein complex responds to glucose, c) how the glucose sensors generate an intracellular signal, and d) the nature of an additional, novel glucose signal transduction pathway that acts on HXT1. To understand the mechanism of glucose repression (Aim2), we need to know a) how Mig1 is regulated by glucose, b) the role of the Reg1-Glc7 protein phosphatase in regulating Mig1 function, and c) the nature of the novel, Snf1-independent mechanism for glucose regulation of the Mig2 repressor. We will also take advantage of structural and functional differences between the glucose sensors and glucose transporters to inform the mechanism of glucose transport across the membrane (Aim 3).